The mechanisms by which proteins sequestered in specific intracellular organelles, but synthesized in the extraorganellar cytoplasm (cytosol), are selectively transported into the organelle where they will reside are presently poorly understood. Recent studies have revealed that a number of such proteins are initially translated as larger precursor molecules which appear to follow one of two different pathways to their receptor organelle. Some of them appear to be made on membrane-bound polysomes and are incorporated into or through organellar membranes co-translationally while others appear to be made on free ribosomes and are incorporated into or through organellar membranes post-translationally. As yet, the molecular determinants which favor one pathway or the other, and the role of transient "leader" sequences in the initial targeting of polypeptides to their receptor membrane for insertion either co-translationally or post-translationally are still elusive. In this study we will examine the mechanisms which underly the sequestion of proteins into organelles by studying the pathways followed by cytoplasmically-translated proteins which reside in the different compartments of yeast mitochondria. By use of antibodies against proteins which reside in different mitochondrial compartments, in vitro translation and transport systems, and "mitochondrial protein transport" defective yeast mutants we hope to: 1) identify cytosolic precursor molecules to mitochondrial proteins; 2) define the ribosomal sites of synthesis and transport pathways followed by precursor molecules during their journey from their site(s) of synthesis to their sites of deposition in the mitochondrion; and 3) examine the protein components of the transport system(s) involved in importing cytosolic precursors into the mitochondrion.